Recent studies using cell culture models of axon regeneration and a biochemical approach have demonstrated that many mRNAs are targeted and translated within regenerating axons following injury. As recent studies have also shown that the local translation of specific mRNAs directly within the growth cones plays a critical role in axon guidance in vitro, an important question is whether specific mRNAs are targeted and translated within the growth cone of regenerating neurons in vitro. It is also critical to extend the above findings from cell culture into in vivo models for nerve regeneration. Almost nothing is known on role of local protein synthesis in vivo to promote axon regeneration. Future work is also critically needed to identify molecular mechanisms involved in mRNA localization and local protein synthesis during development and regeneration. Continued studies to identify underlying mechanisms and their extrapolation in vivo will provide critical tools and approaches to test the hypothesis that specific mRNA localization and translation is necessary for optimal axon regeneration. We have a long-standing interest in studying the mechanism, regulation and function of mRNA transport and local protein synthesis in cultured neurons. An area of technical expertise has been the use of high resolution fluorescence microscopy and digital imaging methods to visualize the localization and dynamic trafficking of mRNAs in fixed and live neurons respectively. We propose to use this expertise to investigate the role of specific mechanisms in mRNA targeting and local translation in axon regeneration, using established in vitro and in vivo models. Aim-1 will use quantitative fluorescence in situ hybridization methods to define whether [unreadable]-actin mRNAs and others are localized specifically within the growth cone of regenerating axons. Live cell imaging methods will be applied to visualize mRNA transport in the axon and its translation at the growth cone. Knockdown approaches and overexpression approaches will be used to assess whether specific mRNA binding proteins can influence axon regeneration. Aim-2 will involve the use of an established in vivo model to study axon regeneration which will be used, in similar fashion to aim-1, to document the presence of specific mRNAs, mRNA binding proteins and translational components, followed by efforts to manipulate the molecular mechanisms with viral vectors and affect axon regeneration in vivo. The proposed studies will provide new insight into novel mechanisms for axon regeneration which will suggest new targets for therapeutic targets for treatment of nerve injury. Axonal degeneration is a common pathway leading to loss of neural function, as seen in various neurological diseases and after spinal cord and nerve injuries. Therapeutic approaches to enhance axon regeneration are very limited. This proposal will investigate a role for specific mRNA targeting and translation mechanisms in optimal nerve regeneration. We anticipate that the proposed basic research will provide new insight into molecular mechanisms underlying optimal axon regeneration and has important implications for future developments of therapies and treatments for nerve injury. [unreadable] [unreadable]